The present invention relates generally to a liquid flow system containing suspended biological cells and particularly to such a system which permits automatic detection of the cells and orientation thereof with respect to the axis of the liquid flow.
Liquid flow systems containing suspended particles such as biological cells are well known in the art at this time. An early flow system of this type is described in one of the Applicants' prior U.S. Pat. No. 3,675,768 and a division thereof U.S. Pat. No. 3,770,349. Since the time of the above referred to United States patents automatic detection of cells for cancer diagnosis and other research programs has become of increasing importance. However, certain problems remain to be solved. Among these problems is the orientation of a cell in the liquid stream when the cell is not spherical.
For the automatic analysis of biological cells suspended in a liquid it is desirable to separate the cells so they flow individually through the liquid. In the past, it has been attempted to disrupt the cytoplasma of the cell to obtain the nucleus of the cell. Blood cells which occur naturally isolated from each other have had the greatest success in liquid flow systems. However, when it is desirable to analyze epithelial cells such as those exfoliated from vaginal epithelia, they must first be processed so as to separate those cells which naturally exfoliate as single or individual cells and to eliminate clumps or clusters. A number of techniques are known for isolating single cells from clumps of cells. These techniques require chemical methods such as treatment with enzymes such as collagenase or hyalorunidase. Also physical methods are known for this purpose such as stirring, irradiating the cells with ultrasound or syringing them. Frequently both physical and chemical methods are required to loosen the cell boundaries.
The preparation of samples for automatic cell analysis may, for example, be described in connection with a collection of cells for cancer detection. Sputum, scrapings from oral mucosa or from the vaginal canal including the cervical opening and the endocervical canal are a complex mixture of epithelial cells which may be rod-shaped, or disk-shaped and may include non-epithelial cells such as spherical leucocytes and disk-like erythrocytes. The size and thickness of the cells depends on the status of the cell differentiation. Most of these cells have a nucleus that contains the most important information with regard of whether the cell is cancerous or not. Also among the cell collections there may be bacterial parasites and mucus. As mentioned before, some cells occur isolated from others while other cells remain attached to each other by their loosened borders; still others occur in clusters which first must be loosened or removed to obtain a suspension of single cells.
Routinely and conventionally the cell samples must be preserved in some manner before the automatic examination can take place. Usually the preservation is accomplished by a fixative chemical such as formaldehyde or ethyl alcohol which coagulates colloids such as proteins. Fixatives and other chemicals change the physical properties of the cell membranes and cause them to be easily permeable by aqueous solutions. Preservation may also be accomplished by other means, for example by refrigeration.
Preservation is required because the samples are taken either at bedside of the patient or in the doctor's office. On the other hand, the cell analysis of all collected samples is usually performed at a different place. Most of the techniques for the isolation of cells are performed on fresh samples before the colloids coagulate. Thus, preferably, the sample preparation is accomplished as close as possible to the time the sample is taken.
For example, most of the uterine or sputum cells to be investigated have the shape of a disk, each cell having a disk-like nucleus. When such cells are attached to a glass surface usually one of the cell's major surfaces sticks to the glass. Because the liquid flow creates hydraulic forces, the cells move parallel to the flow. Hence, for optical image analysis the cell should pass through the focal plane of a microscope. This may or may not be true of cells having a non-spherical shape. If the flow is parallel to the optical axis, the cells are in the focal plane at one instance. However, the nucleus will not present a front image unless the cell has been oriented so that it crosses the focal plane with its major surface parallel to the optical axis. In the prior art this goal has not been attained.
It is accordingly an object of the present invention to prepare biological cells for automatic analysis by separating them from a cell cluster and flattening wrinkled cells.
Another general object of the invention is to control the cells in a liquid flow system by orienting the cells in a desired direction, centering the cells to improve the detection geometry, deflecting selected cells by induction to focus them and spread them in space according to their mass to facilitate sorting of the cells.